For malaria to be spread from person to person via a mosquito, Plasmodium parasites must undergo sexual differentiation to form gametocytes. Although an essential part of the life cycle, little is known about the production of these stages n vitro or in vivo, which complicates the development of strategies that effectively block transmission. In the previous funding period we identified a gene that is critical for gametocyte production, P. falciparum gametocyte development 1 (Pfgdv1) and the set of genes specifically expressed during early gametocytogenesis in P. falciparum (Pfge genes). Analysis of the expression profiles of these genes in vitro and in a cohort of malaria infected patients lead to th hypothesis that gametocytes are formed during each asexual cycle as part of normal development. This type of continuous gametocyte production would provide a consistent source of infectious parasites whenever a mosquito bites. However, it also has serious implications for the design of control measures and suggests that mass treatment or vaccination would be needed to eliminate the parasite. To further evaluate the initiation of gametocytogenesis in vivo, we developed a method to directly compare asexual parasitemia and gametocyte commitment in blood samples from patients. The relationship between gametocyte induction and maturation in vivo is needed to understand the factors that contribute to the production of infectious gametocytes. Aim one will evaluate the role of patient age and hematocrit in gametocyte production, while Aims 2 and 3 will use molecular (Aim 2) and immunological (Aim 3) to understand the role of parasite exposure and immune stimulation on gametocyte commitment and maturation. This field work will complement our ongoing basic research defining the molecular basis for gametocytogenesis. Together the work will extend our understanding of the initiation and formation gametocytes that are essential for the spread of malaria. The findings should provide markers to identify gametocyte carriers before they are infectious and identify signaling pathways that could be targeted to block transmission.